1. Field of the Invention
The present invention relates to 2,4,5-trichlorophenoxyalkyl amide derivatives and an immunoassay of dioxins using the same.
2. Description of the Related Art
The term “dioxins” is a general term for polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs) and coplanar polychlorinated biphenyl (Co-PCBs). For PCDDs and PCDFs, there are a very large number of isomers, such as 75 and 135 isomers, respectively, depending on the number and the position of chlorines, and the intensity of toxicity is considerably different from isomer to isomer. In particular, in PCDD and PCDF isomers, isomers having chlorine at the 2-, 3-, 7- and 8-positions are highly toxic. Examples of the toxic symptoms include thymus atrophy, liver hypertrophy, exhaustion and starvation symptoms, and chloric acne. Furthermore, it is known that long exposure to dioxin, even though only in a low amount, causes chronic symptoms such as porphyria cutanea tarda, and exhibits a variety of toxicity such as teratogenicity, carcinogenicity and cocarcinogenicity.
Furthermore, in recent years, it has become known that a certain type of compounds has the effect of disturbing the endocrine function of organisms, and these compounds, which are so-called “endocrine disturbing chemicals”, have gained attention as a global environmental problem. It also has been found that the dioxins are possible endocrine disturbing chemicals having estrogen activity.
It has become evident that dioxins having such various toxicities are contained in chemical products such as herbicides or insecticides, exhaust gas and fly ash released from incineration plants, or drains from paper mills. For this reason, dioxins are detected not only in samples derived from environments such as water, bottom sediments of rivers or harbors around big cities, air, soil or the like, but also in samples derived from organisms such as food, blood, mother milk, or urine, and pollution is wide-spread, which is regarded as a problem. Thus, since the dioxins in the environment cause a large social problem, it is a pressing task to determine the amount of the dioxins present in the environment.
For determining the dioxins, analytical values with a high precision are required, so that in general, the determination of the dioxins is performed according to an official method of analysis. In this analysis method, various types of chromatography used for extraction, concentration, purification or the like, and equipment including an expensive analysis device such as a high resolution gas chromatograph mass spectrometer are used. The official method of analysis is highly sensitive and can perform multicomponent analysis in which a plurality of compounds can be identified and determined quantitatively at a time. However, this method requires expensive and special equipment and a facility such as a clean room, and also requires a skilled technique for the analysis. Furthermore, it takes time to obtain results because of the complexity of the pre-treatment. Therefore, there is a demand for development of a highly precise and simple determining method. In order to solve this problem, there is a demand for a method for determining dioxins by an immunoassay utilizing a comparatively inexpensive and simple antigen-antibody reaction.
The immunoassay is a method for detecting a trace amount of an antigen using the ability of an antibody to recognize an antigen specifically, and the antigen can be determined with high sensitivity by utilizing the high affinity and the high specificity of the antibody to the antigen. The pre-treatment of samples is simple, so that multiple samples can be determined simply and rapidly, and the cost for the determination is low. Thus, this method is advantageous, so that it is widely used for medicine, biochemistry, pharmacology, agriculture and the like.
It is necessary to recognize an antibody or an antigen in order to detect a substance to be determined in the immunoassay, and various labels are used. Examples of the immunoassay include enzyme immunoassay (EIA), radioimmunoassay (RIA), and fluorescence immunoassay (FIA), which are defined depending on the type of the label. In all of these immunoassays, a calibration curve is obtained by performing an assay with the same compound as a compound to be determined in the same manner as for a sample, and the compound concentration in the sample is calculated based on this calibration curve.
In general, in the determination in clinical tests or the biochemical field, the EIA is utilized because of its simplicity, and the EIA is also attempted to be used for the determination of the dioxins. However, for the dioxins, there are three types having a specific basic structure, as described above. There are a large number of isomers having a different number of chlorine substituents. Therefore, there is a problem as to which compound should be used as the indicator. For example, many of the developed EIA determination systems of dioxins is aimed to determine 2,3,7,8-tetrachlorozibenzo-p-dioxin (2,3,7,8-TeCDD), which is the most toxic substance among the dioxins. Using this toxic substance as a standard, determination of the dioxin has been performed (see Anal. Chem., vol.70, pp. 1092–1099).
However, in the above described assays, since the antigens, such as labeled antigens or solid-phase antigens, are prepared by highly toxic substances, carefully handling and disposal of these toxic antigens is required, which restricts the use of the assay.
Therefore, if a compound other than a toxic compound can be used as an antigen, handling can be easy and a safe and rapid assay for dioxins can be provided. Japanese Laid-Open Patent Publications Nos. 2002-128731 and 2002-131316 propose compounds alternative to PCB, but no compound specific to dioxins has been provided yet.